Investigations on the 4-quinolone-3-carboxylic acid motif part 5: modulation of the physicochemical profile of a set of potent and selective cannabinoid-2 receptor ligands through a bioisosteric approach

Abstract

Three heterocyclic systems were selected as potential bioisosteres of the amide linker for a series of 1,6-disubstituted-4-quinolone-3-carboxamides, which are potent and selective CB2 ligands that exhibit poor water solubility, with the aim of improving their physicochemical profile and also of clarifying properties of importance for amide bond mimicry. Among the newly synthesized compounds, a 1,2,3-triazole derivative (1-(adamantan-1-yl)-4-[6-(furan-2-yl)-1,4-dihydro-4-oxo-1-pentylquinolin-3-yl]-1H-1,2,3-triazole) emerged as the most promising in terms of both physicochemical and pharmacodynamic properties. When assayed in vitro, this derivative exhibited inverse agonist activity, whereas, in the formalin test in mice, it produced analgesic effects antagonized by a well-established inverse agonist. Metabolic studies allowed the identification of a side chain hydroxylated derivative as its only metabolite, which, in its racemic form, still showed appreciable CB2 selectivity, but was 150-fold less potent than the parent compound.

Figure 1

Structure of prototypical 4-quinolone-3-carboxamides 1–4, 14, and 17.

Figure 2

Structure of 4-quinolone-3-carboxamide bioisosteres 5–13 and 15–16.

Figure 3

The effect of compounds 11 (diamonds) and 12 (triangles) on [³⁵S]GTPγS binding to hCB₂-CHO cell membranes (n=8). Each symbol represents the mean percentage change in [³⁵S]GTPγS binding ± SEM.

Figure 4

Alternative conformations for compound 12 and 13. Red arrows indicate H-bond acceptor sites.

Figure 5

Effect of compound 11 (1–3 mg/kg, i.p.) alone (a) or in combination, at the dose of 3 mg/kg, with AM630 (1 mg/kg, i.p.) (b) in the formalin test in mice. The total time of the nociceptive response was measured every 5 min and expressed as the total time of the nociceptive responses in min. Results are mean ± SEM (n=8–10 for each group). In panel a, squares represent formalin 1.25%, circles compound 11 1mg/kg, triangles compound 11 3 mg/kg; filled circles and triangles denote statistically significant differences vs formalin. In panel b, squares represent formalin 1.25%, triangles compound 11 3 mg/kg, diamonds AM630 1 mg/kg + compound 11 3 mg/kg; filled triangles denote statistically significant differences (P < 0.05) vs. formalin, while filled diamonds denote statistically significant differences (P < 0.05) vs. 11. One-way analysis of variance followed by a Turkey-Kramer multiple comparisons test was used for data analysis.

Figure 6

Effect of compound 12 (1–3 mg/kg, i.p.) alone (a) or in combination, at the dose of 3 mg/kg, with AM630 (1 mg/kg, i.p.) (b) in the formalin test in mice. The total time of the nociceptive response was measured every 5 min and expressed as the total time of the nociceptive responses in min. Results are mean ± SEM (n=8–10 for each group). In panel a, squares represent formalin 1.25%, circles compound 12 1mg/kg, triangles compound 12 3 mg/kg; filled circles and triangles denote statistically significant differences vs formalin. In panel b, squares represent formalin 1.25%, triangles compound 12 3 mg/kg, diamonds AM630 1 mg/kg + compound 12 3 mg/kg; filled triangles denote statistically significant differences (P < 0.05) vs. formalin, while filled diamonds denote statistically significant differences (P < 0.05) vs. 12. One-way analysis of variance followed by a Turkey-Kramer multiple comparisons test was used for data analysis.

Figure 7

HPLC-MS analysis of compound 32 and of the CYP-depend metabolite of compound 11. (A) HPLC (total ion current) profile of the reaction mixture obtained by incubating compound 11 with human microsomal preparations (B) Chromatogram of the peak at 499 m/z present in the human microsomal incubation mixture. (C) Chromatogram of the peak at 499 m/z of authentic compound 32.

Figure 8

MS spectra of authentic compound 32 (panel A) and of the CYP-dependent metabolite of compound 11 (panel B).

Scheme 1

Synthesis of compounds 5, 8, 11. Reagents and conditions. a) BMICl, H₂O, MW, 240 °C, sealed tube. b) i. HMT, TFA, MW, 120 °C, sealed tube; ii. H₂O. c) i.dimethyl-1-diazo-2-oxopropylphosphonate, K₂CO₃, MeOH/THF 1/1; ii. AdN₃, CuI, MW, 100 °C, sealed tube. d) R-B(OH)₂, Pd(OAc)₂, PPh₃, 1N Na₂CO₃, DME, EtOH, MW, 150 °C.

Scheme 2

Synthesis of compounds 6, 9, 12. Reagents and conditions. a) NH₂NH₂.H₂O, EtOH, reflux. b) 1-adamantanecarbonyl chloride, DMAP, Et₃N, CH₂Cl₂, 0 °C→rt. c) POCl₃, 110 °C. d) R-B(OH)₂, Pd(OAc)₂, PPh₃, 1N Na₂CO₃, DME, EtOH, MW, 150 °C.

Scheme 3

Synthesis of compounds 7, 10, 13. Reagents and conditions. a) 1-adamantanecarboxamidoxime, HBTU, DIPEA, DMF. b) MW, 150 °C, 10 min. c) R-B(OH)₂, Pd(OAc)₂, PPh₃, 1N Na₂CO₃, DME, EtOH, MW, 150 °C.

Scheme 4

Synthesis of compounds 15 and 16. Reagents and conditions. a) pentyl iodide, K₂CO₃, 18-crown-6, CH₃CN, 90 °C. b) i. TMSA, CuI, Pd(Ph₃)₂Cl₂, DIPEA, dioxane, MW, 120 °C, sealed tube; ii. K₂CO₃, MeOH, THF. c) AdN₃, CuSO₄, sodium ascorbate, MW, 150 °C, sealed tube, DMF.

Scheme 5

Synthesis of compounds 29 and 32. Reagents and conditions. a) ethyl acrylate, TRITON B, DMF, 90 °C. b) TMSA, CuI, Pd(Ph₃)₂Cl₂, DIPEA, THF. c) KF, EtOH. d) AdN₃, CuSO₄, sodium ascorbate, tBuOH/H₂O, 1/1. e) 2-furyl boronic acid, Pd(OA c)₂, PPh₃, 1N Na₂CO₃, DME, EtOH, MW, 150 °C. f) i. (5-iodopentan-2-yloxy)(tert-butyl)dimethylsilane, K₂CO₃, DMF, 90 °C;. ii. TBAF, THF.